A method for extracting L-methionine from fermentation broth

A technology of methionine and fermented liquid, applied in the field of biochemical engineering, can solve problems such as increasing energy consumption

Active Publication Date: 2020-10-27
ZHEJIANG UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In this method, the fermented liquid needs to be heated to a very high temperature, which greatly increases energy consumption.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] 1. Methionine fermentation broth

[0027] Step 1: The metJ and metI genes in E. coli E.coliW3110 were knocked out and then introduced into the pTrc99A / metA* / yjeH plasmid to construct E.coliW3110ΔmetJΔmetI / pTrc99A / metA* / yjeH (Jiangfeng Huang et al.2016.Metabolic engineering of Escherichia coli for microbial production of L-methionine. Biotechnology and Bioengineering. 114:843-851). The E.coliW3110ΔmetJΔmetI / pTrc99A / metA* / yjeH strain was first inoculated into 100mL LB medium, and cultivated overnight on a shaker at a temperature of 37°C and a rotation speed of 200rpm.

[0028] Step 2: Then inoculate 100mL of the culture solution in step 1 into a fermenter with 3.0L of MS medium for culture, and culture and ferment at 30°C and 150rpm until OD600=0.8-1.0, add IPTG with a final concentration of 0.1mM, Continue to cultivate to 48h. Obtained 3.0L fermented broth, liquid phase detection L-methionine content 16.4g / L.

[0029] LB medium: 10.0g / L peptone, 5.0g / L yeast extract, ...

Embodiment 2

[0038] 1. Methionine fermented liquid, same as embodiment 1.

[0039] 2. Extraction of L-methionine

[0040]Step 1: Take 3.0L of L-methionine fermentation broth, adjust the pH of the fermentation broth to 1.5, and then filter it with a ceramic membrane (pore size 50nm) to remove bacterial protein to obtain 2.9L of pretreatment solution.

[0041] Step 2: Get 100mL pretreatment solution and add the hydrogen type D061 cation resin ion exchange column (resin packing column volume is 45mL, high 45cm, ion exchange column aspect ratio is 15: 1). The sample loading flow rate is 1.0BV / h. After sample loading, wash 3.0BV with deionized water to remove some impurities that are not adsorbed on the resin, and then use 2.0mol / L ammonia water to elute at a flow rate of 1.0BV / h. Every 10mL Sampling uses ninhydrin to carry out color reaction (take a drop of effluent and drop it on the filter paper, then drop a drop of 2% ninhydrin solution, dry it, stop collecting if it does not show purple)...

Embodiment 3

[0046] 1. Methionine fermented liquid, same as embodiment 1.

[0047] 2. Extraction of L-methionine

[0048] Step 1: Take 3.0L of L-methionine fermentation broth, adjust the pH of the fermentation broth to 3.0, and then filter it with a ceramic membrane (pore size 50nm) to remove bacterial proteins to obtain 2.9L of pretreatment solution.

[0049] Step 2: Get 100mL pretreatment solution and add the hydrogen type 001x7 cation resin ion exchange column (resin packing column volume is 45mL, high 45cm, ion exchange column aspect ratio is 15: 1). The sample loading flow rate is 1.0BV / h. After sample loading, wash 3.0BV with deionized water to remove some impurities that are not adsorbed on the resin, and then use 2.0mol / L ammonia water to elute at a flow rate of 1.0BV / h. Every 10mL Sampling uses ninhydrin to carry out color reaction (take a drop of effluent and drop it on the filter paper, then drop a drop of 2% ninhydrin solution, dry it, stop collecting if it does not show purp...

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PUM

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Abstract

The invention discloses a method for extracting L-methionine from fermentation liquid. According to the method, regulating the pH of fermentation liquid comprisingcontaining L-methionine to 1.5 to 4;performing filtering; taking filter liquid; using strong acid type cation exchange resin for column chromatography; performing reduced pressure concentration on the eluent comprisingcontaining L-methionine; taking concentrated liquid; adding isopropanol for stirring and crystallization; taking crystals; washing the crystals by ultrapure water until no alcohol exists; performing drying to obtain the L-methionine. The ion exchange resin is used for the first time for separating the methionine from the fermentation liquid; the separation cost is greatly reduced; the yield and the purity can reachthe industrial production level. The method has the advantages that the operation is simple; the steps are short; the used reagent has low cost and can be easily obtained; the relatively good industrialization prospects are realized. The product obtaining yield in the whole process is high; the quality of the finally obtained product is high; the purity is as high as 99 percent; the mass yield isas high as 85 percent.

Description

technical field [0001] The invention relates to the field of biochemical engineering, in particular to a method for extracting L-methionine from fermentation broth. Background technique [0002] Methionine is one of the essential amino acids that constitute the human body and participates in protein synthesis. Because it cannot be produced in the body itself, it must be obtained from the outside. [0003] At present, the method of industrial production of methionine is mainly chemical method. The chemical process uses acrolein and methyl mercaptan as raw materials to produce methionine (TPMA), and then TPMA is combined with sodium cyanide or hydrogen cyanide to synthesize hydantoin. Hydantoin is hydrolyzed by alkali, and then acid ( or CO2) acidification to produce solid methionine, but chemical methods usually only produce DL-methionine. Patent CN104755625A reports a method for chemically producing L-methionine, O-phospho-L-homoserine and methyl mercaptan are converted i...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07C319/28C07C323/58
CPCC07B2200/07C07C319/28C07C323/58
Inventor 柳志强熊能陈涛余茹薛亚平郑裕国
Owner ZHEJIANG UNIV OF TECH
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